HPLC methods for choloroquine determination in biological samples and pharmaceutical products.

OBJECTIVE: Review and assess pharmaceutical and clinical characteristics of chloroquine including high-performance liquid chromatography (HPLC)-based methods used to quantify the drug in pharmaceutical products and biological samples. EVIDENCE ACQUISITION: A literature review was undertaken on the PubMed, Science Direct, and Scielo databases using the following keywords related to the investigated subject: 'chloroquine', 'analytical methods', and 'HPLC'. RESULTS: For more than seven decades, chloroquine has been used to treat malaria and some autoimmune diseases, such as lupus erythematosus and rheumatoid arthritis. There is growing interest in chloroquine as a therapeutic alternative in the treatment of HIV, Q fever, Whipple's disease, fungal, Zika, Chikungunya infections, Sjogren's syndrome, porphyria, chronic ulcerative stomatitis, polymorphic light eruption, and different types of cancer. HPLC coupled to UV detectors is the most employed method to quantify chloroquine in pharmaceutical products and biological samples. The main chromatographic conditions used to identify and quantify chloroquine from tablets and injections, degradation products, and metabolites are presented and discussed. CONCLUSION: Research findings reported in this article may facilitate the repositioning, quality control, and biological monitoring of chloroquine in modern pharmaceutical dosage forms and treatments.

Development and Evaluation of Photoprotective O/W Emulsions Containing Hydroalcoholic Extract of Neoglaziovia variegata (Bromeliaceae).

Neoglaziovia variegata is a Bromeliaceae plant species widely found in Brazil with several pharmacological properties, including photoprotective activity. Although herbal-based active ingredients have been applied in cosmetic products, especially for skin treatment, its application in sunscreen formulations remains unexplored. The aim of this work is to evaluate the photoprotective effect of cosmetic formulations containing hydroalcoholic extract of N. variegata (Nv-HA). Initially, the phenolic and flavonoid total content of Nv-HA were determined. The photoprotective activity of Nv-HA was subsequently assessed using a spectrophotometric method. Nv-HA was incorporated in O/W emulsions in the presence or absence of synthetic filters and their photoprotective efficacy was evaluated by spectrophotometric SPF determination. Finally, the stability study of the formulations was performed through the freezing/defrosting method. Nv-HA showed significant phenolic and flavonoids content (61.66 ± 5.14 mg GAE/g and 90.27 ± 5.03 mg CE/g, resp.). Nv-HA showed SPF values of 5.43 ± 0.07 and 11.73 ± 0.04 for the concentrations of 0.5 and 1.0% (v/v), respectively. It was verified that Nv-HA potentiated the photoprotective effect of formulations containing only synthetic filters. Furthermore, the formulations have remained stable at the end of the preliminary stability study. According to the results, it was concluded that Nv-HA can be used as a chemical filter in cosmetic formulations.

Anti-inflammatory and healing action of oral gel containing borneol monoterpene in chemotherapy-induced mucositis in rats ( Rattus norvegicus )

ABSTRACT The aim of this study was to investigate the effect of gels containing the monoterpene borneol in induced oral mucositis using an animal model. Gels were prepared with borneol at 1.2% and 2.4% (w/w). Oral mucositis was induced by administration of three doses of 5-fluorouracil (30 mg/kg, i.p.) and injury with acetic acid (50%, v/v) soaked in filter paper applied to right cheek mucosa for 60s. Four subgroups comprising 12 animals each were formed. Six animals from each group were sacrificed at days seven and fourteen after oral mucositis induction. Mucous samples were processed and stained with hematoxylin-eosin and Masson's Trichrome. The semiquantitative evaluation involved observation of inflammatory parameters. ImageJ® software was used in the quantitative evaluation. For statistical analyses, Two-way ANOVA, followed by Tukey's post-test (p <0.05), were employed. Borneol 2.4% gel proved effective in the treatment of oral mucositis with statistically significant differences between groups for angiogenesis control, inflammatory cell count reduction and percentage neoformed collagen increase. The confirmation of anti-inflammatory and healing action of borneol in oral mucositis in rats renders it a good marker for predicting this activity for plant extracts rich in this substance

Comparative in vitro performance of three small-volume valved holding chambers with beclomethasone/formoterol pressurized metered dose inhaler.

BACKGROUND: The use of valved holding chambers (VHCs) with pressurized metered dose inhalers (pMDIs) is reported to reduce the oral deposition of inhaled drugs and to facilitate the handling of these devices by patients, especially children. Although the number of commercially available VHCs is increasing, the correct choice of VHC in clinical practice is important, because VHCs are not equally effective regarding medication delivery. Hence, we aimed to evaluate the use of three small-volume VHCs-Vortex(®), AeroChamber(®) Plus (ACP), and Able Spacer™ (AS)-along with a commercial pMDI containing a combination of beclomethasone and formoterol (Innovair(®)) frequently used by asthma patients. METHODS: Evaluation of the delivered dose of both drugs and analysis of particle size distribution of aerosols emitted for the inhaler were performed using the Next Generation Impactor with and without the tested VHCs. RESULTS: The VHCs retained significant quantities of both drugs and dramatically reduced the quantity of drugs deposited in the throat of the impactor, indicating that particles with large size were preferably retained in the VHCs. Interestingly, although the delivered dose of both drugs was reduced by the use of VHCs, the use of the Vortex and the ACP resulted in comparable fine particle doses (FPDs) to that obtained when the pMDI was used alone, whereas the AS VHC significantly reduced the FPDs of both drugs. This may be due to the fact that, unlike the AS VHC, the Vortex and the ACP VHCs are made of antistatic materials that minimize the electrostatic interaction with emitted aerosols, enhancing medication delivery. CONCLUSION: The Vortex and the ACP VHCs present interesting advantages over the AS VHC to be used with Innovair pMDI. However, these results are based on an in vitro evaluation and need to be validated in an in vivo study in order to clinically assess the performance of these VHCs.

Relative bioavailability of two formulations of venlafaxine extended-release 75-mg capsules in healthy brazilian male volunteers: A single-dose, randomized-sequence, open-label, two-period crossover study in the fasting and fed states.

BACKGROUND: The oral antidepressant venlafaxine hydrochloride is a selective serotonin-norepinephrine reuptake inhibitor. OBJECTIVE: The aim of this study was to evaluate the bioequivalence of a new generic formulation of venlafaxine extended-release 75-mg capsules (test) and the available branded formulation (reference) to comply with regulatory criteria for marketing of the test product in Brazil. METHODS: This single-dose, randomized-sequence, open-label, 2-period crossover study was conducted in healthy male volunteers and consisted of separate fast- ing and fed phases. A single oral dose of the test or reference formulation was followed by a 7-day washout period, after which subjects received the alternative formulation. There was a 3-month interval between the fasting and fed portions of the study. There was no standardization of race because of the difficulty of achieving standardization in the Brazilian population. Blood samples were collected before dosing and at 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 5, 6, 8, 10, 12, 24, 36, and 48 hours after dosing. Venlafaxine concentrations were determined using an HPLC-MS/MS method. The formulations were considered bioequivalent if the 90% CIs of the geometric mean ratios (test:reference) for C(max) and AUC(0-t) were within the regulatory range of 80% to 125%. Adverse events were monitored through-out the study based on vital signs, laboratory tests, interviews, and spontaneous patient reports. RESULTS: Forty-eight subjects were enrolled in both phases of the study; all 48 subjects completed the fasting phase, and 1 subject withdrew during the fed phase. The mean (SD) age of participants in the fasting and fed phases was 24.96 (5.5) and 24.90 (4.7) years, respectively; their mean weight was 69.65 (9.6) and 71.00 (10.6) kg and their mean height was 172.0 (6.9) and 173.0 (6.6) cm. Under fasting conditions, the arithmetic mean venlafaxine C(max) was 35.705 (23.946) ng/mL for the test formulation and 34.470 (20.639) ng/mL for the reference formulation, with a geometric mean ratio of 1.04. The arithmetic mean AUC(0-t) for the respective formulations was 562.015 (481.875) and 508.509 (439.456) ng · h/mL, with a geometric mean ratio of 1.11. The arithmetic mean T(max) was 6.188 (1.560) and 5.885 (1.648) hours. Under fed conditions, the arithmetic mean venlafaxine C(max) was 42.892 (24.348) ng/mL for the test formulation and 46.275 (23.011) ng/mL for the reference formulation, with a geometric mean ratio of 0.93. The arithmetic mean AUC(0-t) for the respective formulations was 737.218 (603.998) and 682.124 (524.713) ng · h/mL, with a geometric mean ratio of 1.08. The arithmetic mean T(max) was 6.787 (1.769) and 5.957 (1.661) hours. There were no significant increases in venlafaxine C(max), AUC(0-t), or T(max) for either formulation in the fed phase compared with the fasting phase. In both the fasting and fed portions of the study, the 90% CIs for the ratio (test:reference) of log-transformed C(max) (fasting: 93.24-105.93; fed: 84.67-97.85) and AUC(0-t) (fasting: 102.90-116.71; fed: 98.19-114.41) were within the acceptance range for bioequivalence. The most common adverse events (≥ 5% of subjects) in the fasting phase were nausea (46%), diarrhea (29%), headache (29%), vomiting (15%), and colic (6%); the most common adverse events in the fed phase were nausea (15%), headache (13%), and dizziness (9%). CONCLUSION: In this single-dose study in healthy fasting and fed volunteers, the test formulation of venlafaxine extended-release 75-mg capsules met Brazilian regulatory criteria for bioequivalence to the reference formulation.

Determination of ampicillin in human plasma by solid-phase extraction-liquid chromatography-tandem mass spectrometry (SPE-LC-MS/MS) and its use in bioequivalence studies.

A simple, fast, sensitive and selective solid-phase extraction-liquid chromatography-tandem mass spectrometry (SPE-LC-MS/MS) method for the quantitative analysis of ampicillin (CAS 69-53-4) in human plasma was developed using amoxicillin as internal standard, and sample extraction by solid-phase extraction (SPE). Extracts were separated by reversed-phase C18 with aqueous mobile phase (acetonitrile, 80:20, v/v) with 0.1% formic acid. The method was validated and successfully applied in a bioequivalence study of capsules 500 mg of ampicillin. Using a short running time of 2.5 min, the lower limit of quantification (LLOQ) for obtained ampicillin was 0.1 microg/ml for a plasma sample of 250 microl and a recovery of 94.38% +/- 4.05. Bioequivalence between the products was determined by calculating 90% confidence intervals (CI) for the ratio of Cmax, AUC0-t and AUC0-inf values for the test and reference products, which were within the 0.80-1.25 interval proposed by FDA and EMEA. It is concluded that the two formulations are bioequivalent in their rate and extent of absorption, and thus, may be used interchangeably.

Determination of indinavir in human plasma and its use in pharmacokinetic study

We report the development and validation of a new sensitive, accurate and precise HPLC method with ultraviolet detection for the determination of indinavir sulfate (IND) in human plasma and its application to a bioequivalence study of a new generic formulation. The extraction of IND from plasma samples was achieved by using liquid-liquid extraction with a mean recovery of 73.9 percent. The lower limit of quantification was 0.05 µg/mL. Bioequivalence between the products was determined by calculating 90 percent confidence intervals (CI) for the ratio of Cmax, AUC0-t and AUC0-inf values for the test and reference products, within the 0.80-1.25 interval proposed by ANVISA and FDA. Therefore the medications are bioequivalent and inter-exchangeable.

Foi desenvolvido e validado um novo método, sensível, exato e preciso por Cromatografia a Líquido de Alta Eficiência com detecção em UV para a determinação do sulfato de indinavir (IND) em plasma humano. O fármaco foi extraído do plasma através de extração líquido-líquido obtendo recuperação média de 73.9 por cento, com limite inferior de quantificação de 0.05 µg/mL. O método foi aplicado em estudo de farmacocinética para verificar a bioequivalência entre um novo medicamento genérico e seu produto de referência. A bioequivalência entre os produtos foi determinada calculando-se um intervalo de confiança (IC) de 90 por cento para a razão das médias do produto teste e referência compreendidas entre o intervalo de 0.80-1.25 proposto pela ANVISA e O FDA. Os produtos estudados são bioequivalentes e, portanto, intercambiáveis.

Development and validation of a new method for the quantification of norfloxacin by HPLC-UV and its application to a comparative pharmacokinetic study in human volunteers

The development and validation of a simple and accurate method based on HPLC with ultraviolet detection for the quantification of norfloxacin (NFX) in human plasma and its application to a bioequivalence study between two norfloxacin formulations is described. NFX and the internal standard (cyprofloxacin) were extracted from plasma using liquid-liquid extraction. Chromatographic separation of norfloxacin, cyprofloxacin and plasma interferents was achieved with a C-18 column and a mobile phase consisting of 20 mM sodium hydrogen phosphate buffer pH 3.0 and acetonitrile (88:12, v/v) and quantitation was done at 280 nm. The method was linear from 25 to 3000 ng mL-1 (r² > 0.997578), and norfloxacin and cyprofloxacin had an average recovery from plasma of 93.9 percent and 91.2 percent respectively. The RSD of inter-day quality control samples at the lower limit of quantification was less than 15 percent. After a single oral dose (400 mg) of norfloxacin administered to healthy human volunteers using a randomized 2x2 crossover design, pharmacokinetic parameters (AUC0-t, AUC0-00, Cmax, t1/2) were derived from the plasma concentration curves for both formulations. Pharmacokinetic analysis of the data showed that the two formulations were bioequivalent, while no adverse reactions to the drug were observed.

O desenvolvimento e validação de um método simples e preciso por CLAE-UV para quantificação de norfloxacino (NFX) em plasma humano e a sua aplicação a um estudo de bioequivalência entre duas formulações são descritos. NFX e o padrão interno (ciprofloxacino, PI) foram extraídos do plasma através de extração líquido-líquido. A separação cromatográfica do NFX, do PI e dos interferentes do plasma foi realizada com uma coluna C-18 e fase móvel composta de tampão fosfato de sódio 20 mM pH 3,0 e acetonitrila (88/12, v/v) e quantificado em 280 nm. A resposta do detector aos analitos mostrou-se linear entre 25 a 3000 ng mL-1 (r² > 0,997578) e a recuperação média de NFX e PI foi de 93,9 por cento e 91,2 por cento respectivamente. O desvio padrão relativo de amostras analisadas ao nível do limite inferior de quantificação foi menor que 15 por cento. Foi administrada uma dose de NFX (400 mg) por via oral a voluntários humanos em um estudo aberto, aleatório e cruzado 2x2 entre duas formulações. Os parâmetros farmacocinéticos (AUC0-t, AUC0-00, Cmáx, T1/2) foram observados a partir da curva de concentração versus tempo. A análise farmacocinética mostrou que as duas formulações são bioequivalentes entre si. Nenhum efeito adverso foi observado.